Hydrogen sulfide donors and their THERAPEUTIC potential as antipruritics and anti-inflAmmatory in MOUSE dorsal skin

Leandro Rodrigues1, Eduardo Ekundi-Valentim2, Juliana Florenzano1, Simone Teixeira1, Marcelo Muscará1, Soraia Costa1. 1Department of Pharmacology, Institute of Biomedical Sciences, University of S ã o Paulo, Sã o Paulo, 05.508-900, Brazil, 2Institute of Health Sciences, University Agostinho Neto, Luanda, Angola, 3Department of Pharmacology, Institute of Biomedical Sciences, University of Sã o Paulo, Sã o Paulo, 05.508-900, Brazil, 4Department of Pharmacology, Institute of Biomedical Sciences, University of Sã o Paulo, Sã o Paulo, 05.508-900, Brazil, 5Department of Pharmacology, Institute of Biomedical Sciences, University of Sã o Paulo, Sã o Paulo, 05.508-900, Brazil, 6Department of Pharmacology, Institute of Biomedical Sciences, University of Sã o Paulo, Sã o Paulo, 05.508-900, Brazil

INTRODUCTION: Pruritus (itch) is a sensory modality that, similar to pain, acts as a protective mechanism for the organism. Despite advances in anti-pruritic therapy with histamine antagonists, these agents are ineffective in most patients. Recently, the gaseous mediator H2S has been highlighted as an important modulator of inflammatory and nociceptive processing mechanisms, but its role in pruritus is unknown. This study was carried out to investigate the effects of H2S donors Na2S and Lawesson\'s Reagent (LR) in the pruritus and skin inflammation evoked by histamine and compound 48/80 (C48/80) intradermally (i.d.) injected in mouse dorsal skin. We have also evaluated the kinetics of H2S generation in the naïve skin and its enzyme expression; cystathionine-β-synthase (CBS) and cystathionine-γ-lyase (CSE).

METHODS: Under approval of the Institute of Biomedical Sciences Ethics Committee of University of Sao Paulo (no. 33, book 2/2010), male BALB/c mice (25-30 g) were anaesthetised with isoflurane, and either compound 48/80 (C48/80; 100 µg site-1) or histamine (1µmol site-1) alone or in addition to H2S donors, Na2S (1-100 nmol site-1) and LR (3-300 nmol site-1), was intradermally (i.d.) administered in a single (0.05 ml) injection to produce itching, that was measured as a bout of scratching during 30 min. Skin plasma protein extravasation and neutrophil accumulation was assessed in a separate set of animals by the extravascular accumulation of i.v. injected 125I-albumin and increased myeloperoxidase (MPO) activity in the skin, respectively. Data are presented as mean ± SEM. Stats were performed by ANOVA followed by Bonferroni’s test. P<0.05 was taken as significant.

RESULTS: Either C48/80 or histamine significantly increased itching frequency, and lead to a potent (P<0.01) plasma extravasation when compared to its vehicle Tyrode. The pruritus evoked by histamine, but not C48/80, was significantly inhibited by 67-77 % with the co-injection of Na2S (1-3 nmol site-1, respectively; n=5-9) and by 47-63 % following co-injection of LR (3-30 nmol site-1, respectively; n=6). Either co-injection of Na2S (10–100 nmol site-1; n=5-6) or LR (10–300 nmol site-1; n=5-8) significantly, but not dose-dependently, reduced plasma extravasation induced by C48/80 and histamine or increased MPO activity evoked by C48/80 in the mouse skin. Western blot analyzes revealed, for the first time, that CSE and CBS are constitutively expressed in murine naïve skin in parallel with basal production of H2S.

DISCUSSION: We show for the first time that increasing bioavailability of H2S in the murine skin, by local application of H2S donors, exerts a protective role against histamine-induced pruritus and skin inflammation evoked by histamine and C48/80. We suggest that H2S donors might represent a potential therapeutic class for treatment of pruritus associated with skin inflammation. We also show that there has been a regular production of H2S in naïve skin along with constitutive expression of CSE and CBS.

Acknowledgements: Fapesp, Capes and CNPq for financial support. Barreto MAA & Gouveia IM for technical assistance.